With a liquid chromatograph in which a multichannel detector, such as a photo diode array (PDA) detector, is used as the detector, three-dimensional chromatogram data having the three dimensions of time, wavelength and absorbance can be obtained by repeatedly acquiring an absorption spectrum for an eluate from the exit port of a column, with the point in time of the injection of the sample into the mobile phase as the base point. Similarly, with a liquid chromatograph (LC) or gas chromatograph (GC) in which a mass spectrometer is used as the detector, three-dimensional chromatogram data having the three dimensions of time, mass-to-charge ratio and signal intensity can be obtained by repeatedly performing a scan measurement over a predetermined mass-to-charge-ratio range in the mass spectrometer. The following description deals with the case of a liquid chromatograph using a PDA detector, although the case is the same with a chromatograph using a mass spectrometer as the detector.
FIG. 8A is a model diagram showing three-dimensional chromatogram data obtained with the aforementioned liquid chromatograph. By extracting absorbance data at a specific wavelength (e.g. λ0) from the three-dimensional chromatogram data a wavelength chromatogram showing the relationship between the measurement (i.e. retention time) and the absorbance at that specific wavelength as shown in FIG. 8B can be created. Furthermore, by extracting data which show the absorbance at a specific point in time (measurement time) from the three-dimensional chromatogram data, an absorption spectrum showing the relationship between the wavelength and the absorbance at that point in time can be created.
In such a liquid chromatograph, a quantitative analysis of a known target component is normally performed as follows: A wavelength chromatogram at an absorption wavelength corresponding to that target component is created. On this wavelength chromatogram, the beginning point Ts and ending point Te of a peak originating from the target component are located. The area value of that peak is calculated, and the quantitative value is computed by comparing that area value with a previously obtained calibration curve.
There is no problem with such a quantitative determination of a target component if the peak which has appeared on the extracted wavelength chromatogram originates from only that target component. However, a peak is not always composed of only a single component (target component); it is often the case that a signal originating from an impurity unintended by the analysis operator (or more broadly, any component other than the intended one) is superposed on the peak. If the analysis operator performs the quantitative calculation without noticing such a situation, the result of the quantitative determination will be inaccurate. Accordingly, an impurity determination process (or peak purity determination process) for examining whether a peak located on a chromatogram has originated from only the target component or additionally contains an impurity is often performed in advance of the quantitative calculation.
To date, various methods have been proposed and practically used as the impurity determination process for a peak on a chromatogram. However, the actual situation is such that none of the conventional methods is a decisive solution since each method has both advantages and disadvantages.
For example, in the impurity determination method described in Patent Literature 1, the absorption spectrum obtained at each point in time of the measurement is differentiated with respect to wavelength at a maximum (or minimum) absorption wavelength of the target component to calculate a wavelength differential coefficient, and a differential chromatogram showing the temporal change of the wavelength differential coefficient is created. Whether or not a peak originating from the target component on the wavelength chromatogram contains an impurity is judged by determining whether or not a peak waveform similar to the one which appears on a normal chromatogram is observed on the differential chromatogram. This method is excellent in that whether or not an impurity exists can be determined with a high level of reliability by comparatively simple computations. However, in principle, there is the case where an impurity cannot be detected, as will be hereinafter described.
FIGS. 9A-9C show examples of the relationship between the absorption spectrum originating from a target component (solid line) and the absorption spectrum originating from an impurity (broken line).
In the previously described conventional impurity determination method, as shown in FIG. 9A, the wavelength differential coefficient of the absorption spectrum curve of the impurity at wavelength λ0 where the extreme point of the absorption spectrum originating from the target component is located (i.e. the wavelength at which the wavelength differential coefficient is zero) is used for the impurity determination. As shown in FIG. 9A, if the wavelength at which the absorption spectrum of the impurity is maximized does not coincide with wavelength and therefore the spectrum curve has a certain slope at wavelength λ0, the impurity can be detected. However, as shown in FIG. 9B, if both the extreme point of the absorption spectrum originating from the target component and that of the absorption spectrum originating from the impurity appear at the same wavelength, the wavelength differential coefficient of the absorption spectrum curve of the impurity becomes almost zero, so that the impurity cannot be detected.
Furthermore, as shown in FIG. 9C, if the curve of the absorption spectrum originating from the impurity has a low slope (which can be horizontal in an extreme case) in the vicinity of the extreme point of the absorption spectrum originating from the target component, the impurity-originated peak which appears when the differential chromatogram is created may become extremely low and obscured by noise components, so that it will be ultimately impossible to detect the impurity.
In the case where the sample is introduced by a flow injection analysis (FIA) method without using the column and detected with a PDA detector or similar device, the obtained data will also be three-dimensional data having the three dimensions of time, wavelength and absorbance. Such data are practically equivalent to the three-dimensional chromatogram data collected with a liquid chromatograph. Therefore, three-dimensional data collected by the FIA method should also be included with the “three-dimensional chromatogram data” in the present description.